CA2255731A1 - Non-peptide g-csf mimetics - Google Patents

Abstract

Invented are non-peptide G-CSF mimetics. Also invented are substitued 2,5-Diimino-3a,6a-diaryl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazoles, pharmaceutical compositions containing these compounds, and methods of using these compounds as G-CSF mimetics. Also invented are novel processes used in preparing these compounds.

Description

CA 022~731 1998-11-20 Non-Peptide G-CSF Mimetics BACKGROUND OF THE ~VENTION
Granulocyte colony-stimnl:~ing factor (G-CSF) is a glycoprotein secreted by macrophages, fibroblasts, and endothelial cells originally identified by its ability to stim~ te the survival, proliferation, and differentiation in vitro of predominantly neutrophilic granulocytes from bone marrow progenitors. Nicola, N. A., Annu. Rev.
Biochem. (1989) 58:45. The capacity of G-CSF to regulate in vivo granulopoiesis is ~iU~~ Led by animal and clinical studies, which demonstrated a reversible rise in circnl~ting neu~ phil levels in response to ~n ini~tçred recombinant G-CSF.
Gabrilove, J. L. et al., N. En~l. J. Med. (1988) 318:1414. G-CSF has pleiotropiceffects on mature neutrophils, enhancing their survival and stim~ ting functional activation, including induction of neutrophil ~Ik~line phosphatase (Sato. N. et al., J.
Cell. Physiol. (1988) 37:272) arld high affinity IgA Fc Itcep~ (Weisbart, R. H., et al., Nature (Lond.) (1988) 332:647), priming for respiratory burst (Nathan, C. F.
Blood (1989) 73:301) and increased chemotaxis (Wang, J.M., Blood (1988) 72: 1456). G-CSF effects have also been observed on hematopoietic cells that arenot committ~d to the granulocyte lineage, for example, stim-ll~tion of the proliferation on monocytic dir~erentiation in vitro of some myeloid leukernic cells (Geissler, K., J. Immunol. (1989) 143:140) and the proliferation in vitro of some multipotential hematopoietic precursors (Ferrero, D., Blood (1989) 73:402).
Administration of recombinant G-CSF to patients suffering from neullupenia due to various causes in(lir~t~ that G-CSF is beneficial as an adjuvant in chemotherapy and in bone marrow tr~ncpl~nt~tion (Morstyn, G., et al., Trends Pharmacol. Sci. 10, (1989) 154-159). G-CSF activity is also associated with mobilization of hematopoietic stem cells from the marrow to the pt;~ he1dl blood.
(See review article, Good Review article Haylock et al., Blood 89:2233-2258, 1997).
It would be desirable to provide compounds which allow for the tre~tmPnt of neutropenia to enh~nce leukocyte production by acting as a G-CSF mimeti~.
As disclosed herein it has unexpectedly been discovered that certain non-peptide compounds are effective as G-CSF mimetics.

. .

CA 022~731 1998-11-20 Wo 97/44033 PCT/USg7/08864 As disclosed herein it has unexpectedly been discovered that certain selected 2,~-Diimino-3a,6a-diaryl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imifl~7.oles are effective as G-CSF mimetics.

SUMMARY OF THE INVENTION
This invention relates to compounds of Formula (I):
R4-N\ H
)--N Rl R2>~\--H

H N--R3 (r) wherein Rl, R2, R3 and R4 are independently aryl, where aryl is cyclic or polycyclic aromatic C3-C12, optionally cont~ining one or more heteroatoms, provided that when C is 3 the aromatic ring contains atleast two heteroatoms, and when C is 4 the aromatic ring contains at least one heteroatom, and optionally ~b~ "~e~l with one or more substituents selected fromthe group con.cicting of: C(o)NR6R7 and NR6R7, aryloxy, cycloalkyl, ~ub~liluLed cycloalkyl, alkyl, C6-C12aryl, alkoxy, acyloxy, substituted C6-C12aryl, amino, N-acylamino, nitro, cyano, halogen, hydroxy, -C(O)OR6, -S(o)nR5, protected -OH
and alkyl substituted with one or more substituents selected from the group con~i.cting of: alkoxy, acyloxy, C6-C12aryl, substituted C6-C12aryl, amino, N-acylamino, oxo, hydroxy, cycloalkyl, ~ubsliluLed cycloalkyl, -C(O)OR6, -S(o)nR7,aryloxy, nitro, cyano, halogen and protected -OH, where R6 is hydrogen or alkyl, nisO-2, R7 is hydrogen or alkyl and R5 is hydrogen, cycloalkyl, C6-C12aryl, substituted cycloalkyl, s~lbstitllte~l C6-C 12aryl, alkyl or alkyl substituted with one or more substituents selected from the group con~i~ring of: alkoxy, acyloxy, aryloxy, amino, N-acylarnino, oxo, hydroxy, -C(O)OR6, -S(o)nR7, nitro, cyano, cycloalkyl, substituted cycloalkyl, halogen, C6-C12aryl, substitl-ted C6-C12aryl and protected -OH, where R6 is hydrogen or alkyl, n is 0-2 and R7 is hydrogen or alkyl; and ph~rrn~eutically acceptable salts, hydrates, solvates and esters thereof.
The present invention also relates to the discovery that the compounds of Formula (I) are active as G-CSF mim~tiC,c.
The invention also is a method for treating neull~cnia, inf~hl~ling chPmoth~rapy-influce~l neutropenia and bone marrow tr~ncpl~nt~tion and in mobilizing ~eli~he.dl blood stem cells and other conditions with depressed leukocyte production in ~ lc, inclu~ling hllm~c, which coln~lises ~mini.ctering to a subject in need thereof an effective amount of a ~lcselltly invented G-CSF mim-~.tic compound.
The invention is also a method for treating b~cte-i~l and fungal infections in m~mm~l.c, inrlu-ling hllm~ns~ which coln~l;ses ~-lminict~ring to a subject in need thereof an effective amount of a ylesently invented G-CSF mimeti-~ compound In a further aspect of the invention there is provided novel ylucesses useful inple~alidg the ~lcsen~y invented G-CSF mimetic compounds.
Included in the present invention are ph~rrn~ eutic~l compositions co~ lising a ph~ euti-~l carrier and compounds useful in the methods of the invention.
Also inch--led in the present invention are mPthodc of co-a~lminictering the presently invented G-CSF mim~.tic compounds with further active ingredients.
This invention also relates to the discovery that certain non-peptide compounds are effective as G-CSF mim~tirs DETAILED DESCRIPTION OF THE INVENTION
The compounds of this invention that act as G-CSF mim~tics have the following Formula (I):

CA 022~731 1998-11-20 Wo 97/44033 PCT/USg7/08864 R4-N ,H
~R1 R2~ N--H
, N--(~
N--R (I) wherein R l, R2, R3 and R4 are independently aryl, where aryl is cyclic or polycyclic aromatic C3-C12, optionally cont~ining 5 one or more heteroatoms, provided that when C is 3 the aromatic ring contains at least two heteroatoms, and when C is 4 the aromatic ring contains at least one heteroatom, and optionally substituted with one or more substituents selected from the group con~icting of: C(o)NR6R7 and NR6R7, aryloxy, cycloalkyl, substituted cycloalkyl, alkyl, C6-C12aryl, alkoxy, acyloxy, substituted C6-C12aryl, amino, N-10 acylamino, nitro, cyano, halogen, hydroxy, -C(O)OR6, -S(O)nR5, protected -OH
and alkyl substituted with one or more substitn~nt~ selected from the group con~i~ting of: alkoxy, acyloxy, C6-C12aryl, substituted C6-C12aryl, amino, N-acylamino, oxo, hydroxy, cycloalkyl, substituted cycloalkyl, -C(O)OR6, -S(o)nR7,aryloxy, nitro, cyano, halogen and protected -OH,~5 where R6 is hydrogen or alkyl, nisO-2, R7 is hydrogen or alkyl and R5 is hydrogen, cycloalkyl, C6-C12aryl, sul~s~ l cycloalkyl, substituted 20 C6-C12aryl, alkyl or alkyl ~ubsliluled with one or more substin~ent~ selected from the group con~i~ting of: alkoxy, acyloxy, aryloxy, amino, N-acylarnino, oxo, hydroxy, -C(O)OR6, -S(o)nR7, nitro, cyano, cycloalkyl, ~ cycloalkyl, halogen, C6-C12aryl, substituted C6-C12aryl and protected -OH, where R6 is hydrogen or alkyl, n is 0-2 and R7 is hydrogen or alkyl; and~5 pharmaceutically acceptable salts, hydrates, solvates and esters thereof.
Preferred among the presently invented Fromula I compounds are those in which aryl is: Cs-C12aryl, optionally cont~ining one or two heteroatoms and optionally substituted with one or more substituents selected from the group conci.c~ing of: -OC6-C I 2aryl, -(CH2)mOH, C6-C 1 2aryl, C I -C4alkyl,-OC 1 -C4alkyl, amino, nitro, cyano, N-acylarnino, trifluoromethyl, C3 7cycloalkyl, halogen, -(CH2)pCOOH, -S(o)nR7 and protected -OH, where m is 0-4, p is 0-3, n is 0-2 and R7 is hydrogen or C 1 4alkyl; and S ph~rrn~nelltic~lly acceptable salts, hydrates, solvates and esters thereof.
Particularly preferred among the presently invented compounds are those in which R 1 and R2 are independently phenyl, naphthyl, furyl, thienyl, pyridyl, indolyl or quinolyl all of which are un~ub~ uled or substituted with a substituent selected from the group comiC~ing of: halogen, Cl salkyl, C3 7cycloalkyl and -0-C14alkyl;R3 and R4 are indepen-lPntly phenyl, naphthyl, thienyl, pyridyl, quinolyl, ~e~.7.i-..id~7.olyl ben_othi~olyl, ben7.ox~7.olyl, thia_olyl or imi-l~7.olyl all of which are un.cubstitu~ecl or s~lbstitl~tecl with one or two substituents seleGted from the group cnncictin~ of: halogen, amino, cyano, N-acylamino, Cl 3 alkyl, -0-Cl 3 alkyl, nitro, -C02H and CF3; and 15 ph~ el~ically acceptable salts, hydrates, solvates and esters thereof.
Particularly preferred among the presently invented coln~ou"ds are those in which R1 and R2 are indepçn-le.ntly phenyl, furyl, thienyl or pyridyl all of which are un.~uh~lilul~-d or ~ub~liLuled with a substituent selected from the group consisting of:
halogen, C1 salkyl, C3 6cycloalkyl and -0-C1 3alkyl;
R3 and R4 are indepen~ently pyridyl, ben7.i.. id~7.olyl, ben7.othi~7.olyl, ben_oxa_olyl, thiazolyl or imida_olyl all of which are u.,sul,~Liluled or substin-te~l with one or two suhstitu~ntc selectPd from the group con~ictin~ of: halogen, arruno, cyano, N-acylamino, C1 3 alkyl, -0-C1 3 alkyl, nitro, -C02H and CF3; and ph~nn~l~el1tic~11y acceptable salts, hydlales, solvates and esters thereof.
The most ~r~fel,ed compounds of the present invention are those in which R1 and R2 are 2-pyridyl both of which are independently unsubstituted or substituted with C1 3alkyl; and R3 and R4 are 2-ben7.imi~7.01yl both of which are indepe.ll-lently ~ Imcubstitllte-l or substituted with Cl 3alkyl, provided that R3 and R4 are not both substituted.
Preferred arnong the presently invented colllp~ullds are:

.....

CA 022~731 1998-11-20 2,5-bis[2-bçn7imi~701ylirnino]-3a,6a-bis(2-pyridyl)- 1 ,2,3,3a,4,5,6,6a-octahydrnimi~l~7.o[4,5-d]imid~7.ole, 2,5-bis[2-ben7imid~7.olylimino]-3a-(2-pyridyl)-6a-phenyl- 1 ,2,3,3a,4,5,6,6a-octahydroimid~7o[4~s-d]imi~ 7ole~

5-[2-be.-7.i, . .i~1~7.olylirnino]-2-[(4-methyl-2-be~7.imi~1~7.olyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imid~7.ole bis(trifluoro;lcet~e) salt, 5-(2-be..~ 701ylimino)-2-[(S-methyl-2-be..7.;"~ 7.olyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,3,3a,4,5,6,6a-octahydrnimitl~7.o[4,5-d]imic~7.ole bis(trifluoro~rer~rç) salt, 2,5-bis[( l -methyl-2-be n7.; . . ~ 7.olyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydrornidazo[4,5-d]imid~7.ole bis(trifluoro~cet~te) salt, 5-[2-bel 7;n~ 7olylimino]-2-[(s~6-dimethyl-2-ben7imid~7olyl)imino]-3a~6a bis(2-pyridyl)- 1,2,2a,4,5,6,6a-octahydroirnidazo[4,5-d]imid~7.ole lS bis(trifluoroacetate) salt, 2,5-bis[2-be~7.imi~l~701ylimino]-3a,6a-diphenyl- 1 ,2,3,3a,4,5,6,6a-octahydroimida_o[4,5-d]imid~7ole~
2,5-bis[2-b~n7.imid~7.olylimino]-3a,6a-bis(2-furyl)-1,2,3,3a,4,5,6,6a-bis(2-pyridyl)octahydroimidazo[4,5-d]imi~i~7.ole, 2,5-bis[2-bçn7imid~701ylimino]-3a,6a-bis(3-methoxyphenyl)-1,2,3,3a,4,5,6,6a-octahydroimi~l~7.o[4,5-d]imid~7- le7 2,5-bis[2-ben~.;.1.i-1~7olylirnino]-3a,6a-bis(4-methylphenyl)-1,2,3,3a,4,5,6,6a-octahydroimid~7.o[4,5-d]imill~7.ole, 2,5-bis[2-benzimida_olylimino]-3a,6a-bis(4-fluolu~henyl)- 1,2,3 ,3a,4,5,6,6a-octahydroimida_o[4,5-d]imid~7.ol-e, 2,5-bis[2-ben7.imid~7.olylimino]-3a,6a-bis(6-methyl-2-pyridyl)-1 ,2,3,3a,4,5,6,6a-octahydroimi-l~70[4,5-d]imi-l~7.ole, 2,5-bis[2-ben7.imid~701ylimino]-3a-(4-pyridyl)-6a-phenyl- 1,2,3 ,3a,4,5,6,6a-octahydroimi~7.o[4,5-d]imida_ole, 2,5-bis[(S-methyl-2-ben7imirl~7.olyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimitl~7o[4~s-d]im~ 7ole~

CA 022~731 1998-11-20 2,5-bis[(5-nitro-2-bçn7.imid:l701yl)imino]-3a,6a-bis(2-pyridyl)-I ,2,3,3a,4,5,6,6a-octahydroimid~70[4,5-d]imi~1~701e, 2,5-bis[(5-chloro-2-bçn7imiA~7.olyl)imino]-3a,6a-bis(2-pyridyl)-1 ,2,3,3a,4,5,6,6a-octahydroimi-1~7.o[4,5-d]imiA~7.ole, S 2,5-bis[(4-methyl-2-thiazolyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,3,3a,4,5,6a-octahydroimid~7.0[4,5-d]imitl~7.0l~, 2,5-bis[2-benzoxazolylimino]-3a,6a-bis(2-pyridyl)- 1 ,2,3a,4,5,6,6a-octahydroimiA~7.o[4,5-d]imi~1~7~
2,5-bis~(4-methyl-2-bel.~.i."iA~7olyl~imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimiA~7O[4,5-d]imidazole, 5-(2-beh,;...i-1~7olylirnino)-2-[(S-fluoro-2-bel.,.;..,i-1~7.olyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,3,3a,4,5,6,6a-oetahydroimidazo[4,5-dlimidazole bis(trifluor~a~etP~P) salt, 2,5-bis[(5-fluoro-2-bell~.;...iA~701yl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimiA~7.o[4,5-d~imidazole bis(trifluoroaretPt~) salt, 5-(2-bçn7.imi~701ylimino)-2-[(l -methyl-2-ben7.imi~1~7olyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,3,3a,4,5 ,6,6a-oetahydroimiA~7.o[4,5-dlimiA~7ole bis(trifluoro~ret~t~) salt, 5-[2-be~ idazolylimino]-2-[(4-methyl-2-thiazolyl)imino]-3a,6a-bis(2-pyridyl)- 1,2,2a,4,5,6,6a-oetahydroilllidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(5-bromo-4-methyl-2-thiazolyl)imino] -3a,6a-bis(2-pyridyl)-1 ,2,3 ,3a,4,5,6a-octahydroirnidazo[4,5-d~imiA~7-11e, 5-[2-bellzilllidazolylimino]-2-[(5-chloro-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoro~eet~t~) salt, 2,5-bis[(5,6-dimethyl-2-ben7imi-1~7.olyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoro~et~tr) salt, 2,5-bis[(S-carboxy-2-ber 7.imicl~7Qlyl)imino]-3a,6a-bis(2-pyridyl)-- 1,2,2a,4,5,6,6a-octahydroimitl:~7.o[4,5-d]imidazole bis(trifluoroacetate) salt, 5-[2-benzill~idazolylimino]-2-[2-bçn7o~7olylimino]-3a~6a-bis(2-pyridyl) 1,2,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroaret~te) salt, . ,. ~

CA 022~731 1998-11-20 2,5-bis~2-ben_othia_olylimino]-3a,6a-bis(2-pyridyl)- 1 ,2,3a,4,5,6,6a-octahydroimid~o[4~s-d]imid~7Qle~
5-~2-bsn7imi~7.olylimino]-2-[2-benzothia_olylimino]-3a,6a-bis(2-pyridyl)-1,2,3a,4,5,6,6a-octahydroimil1~7.o[4,5-d]imid~ole bis(trifluoroacetate) salt, 5-[(2-be~ irl~7Qlylimino)]-2-[(S-carboxy-2-ben7.imitl~7.olyl)imino]-3a,6a-bis(2-pyridyl)- 1 ,2,3,3a,4,5,6,6a-octahydroimi~7.o[4,5-d]imitl~7.ole bis(trifluoroacetate) salt, 2,5-bis[(5-iodo-2-ben7.imi(1~7olyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimi~7.o[4,5-d]imid~ole bis(trifluulo~c~ e) salt, and 5-[2-bel.,.i... i~701ylimino]-2-[(5-iodo-2-be,llzi.. ,ida_olyl)irnino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroi-..i~7.o[4,5-d]imid~ole bis(trifluoroacetate) sa3t.
2,5-bis[2-b~ n7.i . - ~ 7.olylimino]-3a,6a-bis(4-bromophenyl)- 1,2,3 ,3a,4,5,6,6a-octahydroimi(l~7Q[4~5-d]imi~7ole~
2,5-bis[2-be.l.7.;~ 7.o3ylimino]-3a,6a-bis(4-trifluololllGIllylphenyl)-1~2~3~3a~4~5~6~6a-octahydroimi~1~7o[4~5-d]imi(l~7ole~
2,5-bis[2-bel17.imid~7.nlylimino]-3a-(3-chlolùph~ yl)-6a-phenyl-1,2,3,3a,4,5,6,6a-octahydlui~ 7.o[4~5-d]imid~7.ole~
2,5-bis[2-ben7.imi(1~7.olylimino]-3a-(2-chloruphel,yl)-6a-phenyl-1 ,2,3,3a,4,5,6,6a-octahy-llui . ~ 7.o[4,5-d]imid~7.ole, 5-(2-bel.,.;-.. i-~7.olylimino)-2-[(5-methyl-2-be~7.imid~7.Qlyl)imino]-6a-phenyl-3a-(2-pyridyl)- 1 ,2,3,3a,4,5,6,6a-octahydroimid~o[4,5-d]imid~7.Qle bis(trifluoroacetate) salt, 2,5-bis[(l-methyl-2-be,n~.i---i~1~7.Qlyl)imino]-3a,6a-diphenyl-1,2,3,3a,4,5,6,6a-octahydlulllida_o[4,5-d];mi~l~7.Qle, 2,5-bis[(1-methyl-2-ben7.imi~1~7.olyl)imino]-3a,6a-bis(4-lllc;lhyl~h~l.yl)-1,2,3,3a,4,5,6,6a-octahydromid~o[4,5-d]imid~ole, and 2,5-bis[( 1 -methyl-2-ben7.imi~1~7olyl)imino]-3a,6a-bis(4-brQmophenyl)-1 ,2,3,3a,4,5,6,6a-octahydromida_o[4,5-d]imi(~~.ole.
By the term "protected hydroxy" or "protected -OH" as used herein, is meant 30 the alcoholic or carboxylic-OH groups which can be protected by conventional CA 022~731 1998-11-20 blocking groups in the art as described in "Protective Groups In Organic Synthesis"
by Theodora W. Greene, Wiley-Interscience, 1981, New York.
By the term "Cs-C12 aryl" as used herein, unless otherwise defined, is meant a cyclic or polycyclic aromatic Cs-C12 optionally cont~ining one or two 5 heteroatoms.
By the terrn "C6-C12 aryl" as used herein, unless otherwise ~lefine-l, is meant phenyl, naphthyl, 3,4-methylenedioxyphenyl, pyridyl, or biphenyl.
By the term "substituted" as used herein, unless otherwise defined, is meant that the subject ch~rni~l moiety has one or more substituents selected from the 10 group conCicting of: hydroxyalkyl, alkoxy, acyloxy, alkyl, amino, N-acylamino, hydroxy, -(CH2)gC(O)OR6, -S(o)nR7, nitro, cyano, halogen, trifluoro~ llyl and protected -OH, where g is 0-6, R6 is hydrogen or alkyl, n is 0-2, and R7 is hydrogen or alkyl.
By the term "alkoxy" as used herein is meant -Oalkyl where alkyl is as 15 described herein in~ ling -OCH3 and -OC(CH3)2CH3.
The term "cycloalkyl" as used herein unless otherwise defined, is meant a nonaromatic, ullsdlulated or saturated, cyclic or polycyclic C3-C 12 Examples of cycloalkyl and ~ub~ Jled cycloalkyl substituentc as used herein include: cyclohexyl, 4-hydroxy-cyclohexyl, 2-ethylcyclohexyl, propyl 4-20 methoxycyclohexyl, 4-methoxycyclohexyl, 4-carboxycyclohexyl and cyclopentyl.
By the term "acyloxy" as used herein is meant -OC(O)alkyl where aL~yl is as described herein. F.Y~mples of acyloxy substituents as used herein include: -OC(O)CH3, -OC(O)CH(CH3)2 and -OC(O)(CH2)3CH3.
By the term "N-acylamino" as used herein is meant -N(H)C(O)alkyl, where 25 alkyl is as described herein. F.Y~mplçs of N-acylamino suhstitu~ont.c as used herein include: -N(H)C(O)CH3, -N(H)C(O)CH(CH3)2 and -N(H)C(O)(CH2)3CH3.
By the term "aryloxy" as used herein is meant -OC6-C12aryl where C6-C12aryl is phenyl, naphthyl, 3,4-methylenedioxyphenyl, pyridyl or biphenyl optionally substituted with one or more substituents selected from the group 30 con.ci.cting of: alkyl, hydroxyalkyl, alkoxy, trifuloromethyl, acyloxy, amino, N-acylamino, hydroxy, -(CH2)gC(O)OR6, -S(o)nR7, nitro, cyano, halogen and .

CA 022~731 1998-11-20 WO 97/44033 rCT/US97/08864 protected -OH, where g is 0-6, R6 is hydrogen or alkyl, n is 0-2 and R7 is hydrogen or alkyl. Examples of aryloxy substituentc as used herein include: phenoxy, 4-fluorophenyloxy and biphenyloxy.
By the term "heteroatom" as used herein is meant oxygen, nitrogen or sulfur.
S By the term "halogen" as used herein is meant a substituent selected from bromide, iodide, chloride and fluoride.
By the term "alkyl" and derivatives thereof and in all carbon chains as used herein is meant a linear or br~nrhrd, saturated or unsaturated hydrocarbon chainhaving C 1-C 12 carbon atoms. Examples of alkyl substituents as used herein include:-CH3,-CH2-CH3,-CH2-CH2-CH3,-CH(CH3)2,-C(CH3)3,-(CH2)3-CH3, -CH2-CH(CH3)2 and -CH(CH3)-CH2-CH3, -CH=CH2.
By the term "treating" and derivatives thereof as used herein, is meant prophylatic or thel~eulic therapy.
By the phrase "mobilizing p~,lipheldl blood stem cells" as used herein is meant the mobilization of hematopoietic stem cells from the marrow to the peripheral blood.
By the phrase "non-peptide bifunctional ligand" as used herein is meant a ligand capable of activating the G-CSF receptor by binding to two non-~ cent sites.
By the phrase "non-peptide" as used herein is meant protein or peptide not compri.cing primArily of natural amino acids.
By the term "prim:~rily" as used herein, unless otherwise defined, is meant less than 60% by weight of the naturally occurring amino acid residue.
All publications, including but not limited to patents and patent applications, cited in this spec-ifr~iton are herein incorporated by reference as if each individual publicaiton were specifir~lly and individually inriir~trd to be incol~olated by reference herein as though fully set forth.
Coll,~ounds of Formula (I) are includefl in the ph~rm~reutir~l compositions of the invention and used in the m~.tho~ls of the invention. Where a -COOH or -OH
group is present, ph~rm~ceuti~lly acceptable esters can be employed, for examplemethyl, ethyl, pivaloyloxymethyl, and the like for-COOH, and acetate m~ tr and the like for -OH, and those esters known in the art for modifying solubility or hydrolysis char~ct~ri~tics for use as sust~inp~ release or prodrug form~ tions.
By the term "co-a~lmini~tering" and derivatives thereof as used herein is meant either simlllt~ntoous ~riminictration or any manner of separate sequenti~l~rimini~tration of a G-CSF rnimetic compound, as described herein, and a furtheractive ingredient or ingredients, such as antibacterial agents, antifungal agents as well as agents known to treat neutropenia, including ch~mothPrapy-in(i'~ced neuLlo~nia and bone ~ tr~n~pl~nt~tion and in mobilizing peliphel~l blood stem cells and other conditions with depl~,;,sed leukocyte production. Preferably, if the ~rimini.ctration is not ~imlllt~neous~ the compounds are ;~lminictered in a close time proximity to each other. Further nore, it does not matter if the compounds are aAmini~tered in the same dosage form, e.g. one compound may be ~flminictered topically and another compounds may be arlrnini~t~.red orally.
The novel colll~o~ ds of Formula (I) are ~ p~cd as shown in Scheme I
below provided that the 'R' substituentc do not include any such substitl~ent~ that render inopelaLive the Scheme I process. The 1,2-Diketones of Formula (2) are collJ~ ;ially available or prepared from aryl aldehydes by benzoin conclen~tion acconling to the ploce(lufe described in Ide et al (Organic Reactions vol IV, 269) followed by oxidation of interme~ te alpha-hydroxy ketones as reported by Mancuso et al. (J. Org. Chem. 1979, 44, 4148). The gu~ni~line derivatives of Formula (3) are col~..lle.cially available or can be readily prepared from co.~.. -.. ~;ially available o-phenyl.on~ min~s following the procedure of King et al (J. Chem. Soc. 1948, 1366).
Scheme (I) l'~ lion of Compounds of Formula (I) Co~ Jollnds of Formula (I) R4-N' ,H
~R1 , N--(~
H N--R3 (I) wherein R 1, R2, R3 and R4 are as described in Formula (I) above;
are ~l~p~ed by treating a 1,2-diketone of Formula (2) R1~R2 o (2) whelt;in R 1 and R2 are as described in Formula (I) with one or more guanidine derivatives of Formula (3) \>--NH2 NH2 (3) wherein R5 is R3 or R4, such that when more than one gu~ni~lin~o de~ivalive of Formula (3) is utili7.~, R5 is not the same substituent The reaction is run in asolvent such as benzene or pyridine at reflux with azeolropic removal of water such as by using a Dean-Stark apparatus or, alternatively, in the presence of a base catalyst (such as sodium hydroxide) in a suitable protic solvent (such as m~th~nol, ethanol or water).
ph~rm~reuti~lly acceptable salts, hydrates and solvates are formed when app,opliate by m~tho~c well known to those of skill in the art.
Rec~llse the ph~rmS~ ~ellti~lly active co~ ounds of the present invention are active as G-CSF mim~ti-~s they exhibit theldpeulic utility in treating bacterialinfections, fungal infections, neullol)enia, including chemotherapy-in(luced nc.ll.o~nia and bone marrow tr:~n~pl~nt~tion and in mobili7ing peripheral blood stem cells and other conditions with depressed leukocyte production.
In c~e~e~ inin~ potency as G-CSF mimetics, the following assays were employed:
Luciferase Assay Compounds of the present invention were tested for potency as mim~tirs of the G-CSF receptor in a Luciferase assay such as described in Lamb, et al., Nucleic Acids Research 23: 3283-3289 (1995) and Seidel, et al., Proc. Natl. Acad. Sci.. USA
92: 3041-3045 (1995) by substituting NFS60 cells for the HepG2 cells utilized therein. The NFS60 cells (Holmes, et al., Proc. Natl. Acad. Sci. USA 82: 6687-6691 (1985)) were substituted for the HepG2 cells in the Lamb assay because the NSF60 CA 022~731 1998-11-20 Wo 97/44033 PCT/USg7/08864 cells express endogenous G-CSP receptors closely m~tching the pattern of STAT
(signal tr~ncdllcers and activators of transcription) activation observed in primary murine and human bone marrow cells.
Some of the most preferred compounds of this invention were also active in a 5 CFU-G assay, an ex~mple of which is described in King AG, Talmadge J., Badger AM, Pelus LM. Regulation of colony stiml-l~ting activity production from bone marrow stromal cells by the hematoregulatory peptide, HP-5. Exp. Hematol. 20:223-228, 1992.
The ph~rm~e~ltically active compounds within the scope of this invention 10 are useful as G-CSF mimetirs in m~mm~l~, including humans, in need thereof.
The compounds of Examples 1, 2, 4 - 8, 13, 15 - 18, 20, 25, 26, 30, 32b, 34, 35,37 and 39 showed activation above 150% of control between the conce~ d~ion range of 1 to 32 uM in the luciferase assay.
The compounds of Examples 3, 9 - 12, 14, 19, 21 - 24, 27 - 29, 31, 32a, 33, 36,38 and 40, although not showing an activation above 150% of control between the concentration range of 1 to 32 uM, were believed to show activation of above150% of control at > 32 uM in the luciferase assay.
The present invention therefor provides a method of treating bacteri~l infections, fungal infections, neullù~ellia, including chemotherapy-in~luced 20 n~,ullupel1ia and bone marrow transplantation and in mobilizing peripheral blood stem cells and other conditions with depressed leukocyte production, which colll~l,ses ~mini.ct~.ring a compound of Formula (I):

H~NR'H
H N--R3 (I) whe~in R1, R2, R3 and R4 are independently aryl, where aryl is cyclic or polycyclic aromatic C3-C12, optionally cont~ining - one or more heteroatoms, provided that when C is 3 the aromatic ring contains at CA 022~731 1998-11-20 Wo 97/44033 PCT/USg7/08864 least two hetero~tom~, and when C is 4 the aromatic ring contains at least one heteroatom, and optionally sub~lilul~d with one or more substituents selected from the group consisting of: C(o)NR6R7 and NR6R7, aryloxy, cycloalkyl, substituted cycloalkyl, alkyl, C6-C12aryl, alkoxy, acyloxy, substituted C6-C12aryl, amino, N-5 acylamino, nitro, cyano, halogen, hydroxy, -C(O)OR6, -S(o)nR5, protected -OH
and alkyl substituted with one or more substituents selected from the group con~i~ting of: alkoxy, acyloxy, C6-C12aryl, substituted C6-C12aryl, amino, N-acylamino, oxo, hydroxy, cycloalkyl, subsliluled cycloalkyl, -C(O)OR6, -S(o)nR7,aryloxy, nitro, cyano, halogen and protected -OH, 10 where R6 is hydrogen or alkyl, nisO-2, R7 is hydrogen or alkyl and RS is hydrogen, cycloalkyl, C6-C12aryl, substituted cycloalkyl, ~ubs~ilul~d 15 C6-C12aryl, alkyl or alkyl substit-ltecl with one or more substituentc select~l from the group con~i~ting of: alkoxy, acyloxy, aryloxy, amino, N-acylarnino, oxo, hydroxy, -C(O)OR6, -S(o)nR7, nitro, cyano, cycloalkyl, substituted cycloalkyl, halogen, C6-C12aryl, substituted C6-C12aryl and protected -OH, where R6 is hydrogen or alkyl, n is 0-2 and R7 is hydrogen or alkyl; and 20 ph ~rrn~neutically acceptable salts, hydrates, solvates and esters thereof in a quantity effective to enh~n~e leukocyte pro-luctio~ The compounds of Formula (I) also provide for a method of treating the above in~ ts~ disease states because of their demon~trated ability to act as G-CSF mim~.tics. The drug may be ~rlmini~tered to a patient in need thereof by any conventional route of a~lmini~tration, inf~ln~ling, but 25 not limited to, intravenous, intr~mllsc~ r, oral, subcutaneous, intr~1erm~1, and parenteral.
The ph:lrm~ceutically active compounds of the present invention are incol~G ~ted into convenient dosage forrns such as capsules, tablets, or injectable preparations. Solid or liquid ph~rm~ceutical carriers are employed. Solid carriers 30 include, starch, lactose, calcium sulfate dihydrate, terra alba, sucrose, talc, gelatin, agar, pectin, acacia, m~gn~ m stearate, and stearic acid. Liquid carriers include syrup, peanut oil, olive oil, saline, and water. Similarly, the carrier or diluent may include any prolonged release material, such as glyceryl monostearate or glyceryl distearate, alone or with a wax. The amount of solid carrier varies widely but, preferably, will be from about 25 mg to about l g per dosage unit. When a liquid5 carrier is used, the preparation will be in the forrn of a syrup, elixir, emulsion, soft gelatin capsule, sterile injectable liquid such as an arnpoule, or an aqueous ornonaqueous liquid s~.cper-.cion.
The yl~ eutic~l preparations are made following conventional techniques of a ph~rrn~nel-tir~l chPmict involving mixing, gr~n~ ting, and compressing, when 10 neces.c~ry, for tablet forms, or mixing, filling and dissolving the ingreidents, as appn~pliate, to give the desired oral or parenteral products.
Doses of the presently invented ph~rm~rel1tic~lly active compounds in a pl ~ eutical dosage unit as tlPsçribe(i above will be an efficacious, nontoxic quantity plc;fc~dbly selPstecl from the range of 0.001 - 100 mg/kg of active compound, preferably 0.001 - 50 mg~cg. When treating a human patient in need of a G-CSF mimPtir' the selected dose is ~rlminictered preferably from 1-6 times daily, orally or parenterally. Preferred forms of parenteral ~-lmini.ctration include topically, rectally, tr~ncderrn~1ly~ by injection and continuously by infusion. Oral dosage units for human a(lmini.ctration preferably contain from 0.05 to 3500 mg of active 20 compound. Oral ~1mini.ctration, which uses lower dosages is pre~lred. P~clllel~l ~lmini~tration, at high dosages, however, also can be used when safe and convenient for the patient.
Optimal dosages to be ~lminictered may be readily dl~te~ ed by those skilled in the art, and will vary with the particular G-CSF mim~tiG in use, the 25 strength of the ylep~tion, the mode of a~minictration, and the advancement of the disease condition. Additional factors depending on the particular patient being treated will result in a need to adjust dosages, including patient age, weight, diet, and tirne of ~mi~ictration.
The method of this invention of inducing G-CSF mim~tin activity in 30 ".~....n~lc, in~ ling hllm~nc, colll~lises ~lminiclering to a subject in need of such CA 022~731 1998-11-20 activity an effective G-CSF mimPtir. arnount of a ph~rm~r,eutir~lly active compound of the present invention.
The invention also provides for the use of a compound of Formula (I) in the m~nnf~ctllre of a medicament for use as a G-CSF mimrtic.
The invention also provides for the use of a compound of Formula (I) in the manufacture of a medicament for use in therapy.
The invention also provides for the use of a compound of Formula (I) in the m~nuf~rtllre of a mrrlic~mrnt for use in enhancing leukocyte production.
The invention also provides for the use of a co~llpound of Formula (I) in the 10 m~nllfartllre of a m~,-lir~mP,nt for use in treating b~tP-ri~l and fungal infections.
The invention also provides for a pharmaceutical composition for use as a G-CSF mim.o,tic which compri~es a compound of Formula I and a pharm~reutic~lly acceptable carrier.
The invention also provides for a pharm~reutic~l composition for use in the 15 tre~tmrnt of neuL~pel1ia which comprises a co~ ou"d of Formula I and a ph~rm~reutiç~lly acceptable carrier.
The invention also provides for a ph~rm~ceutir~l composition for use in r,nh~nrin~ leukocyte production which comprises a compound of Formula I and a pharm~reutir~lly acceptable carrier.
The invention also provides for a ph~rm~r.eutir~l co,llposilion for use in treating h~ r~ ;~l infections which comprises a compound of Formula I and a ph~rm~reu~ir~lly acceptable carrier.
The invention also provides for a ph~rm~reutir~l colllposilion for use in treating fungal infections which comprises a colllpoLmd of Formula I and a ph~rm~reutir,~lly acceptable carrier.
The invention also provides for a process for p~illg a ph~rm~ce~ltir~l composition cont~ining a pharm~l~eutic~lly acceptable carrier or diluent and a compound of Formula I which comprises bringing the compound of Formula I into association with the ph~ reutically acceptable carrier or diluent.
No unacceptable toxicological effects are expected when compounds of the invention are aflmini~tered in accordance with the present invention.

.. ..

CA 022~731 1998-ll-20 wo 97/44033 PCT/USg7/08864 In addition, the pharm~reutiç~lly active compounds of the present invention can be co-~lmini~tered with further active ingredients, such as other compounds known to treat bacterial infections, fungal infections, neul-upenia, including chPmoth~rapy-in~luce~l neutropenia and bone marrow transplantation and in mobilizing peripheral blood stem cells and other conditions with depressed leukocyte production, or compounds known to have utility when used in combination with a G-CSF mim~.tic.
Without further elaboration, it is believed that one skilled in the art can, using the preceding description, utilize the present invention to its fullest extent.
The following Examples are, therefore, to be construed as merely illustrative and not a limitation of the scope of the present invention in any way.

E~yç. hllf l~t~l Details F.xarr~le 1 Preparation of 2.5-Bisr2-bel~7.i,~ 7.olylirninol-3a.6a-bis(2-pyridyl)-1-2-~.3a.4.5.6.6a-octahydroimidazor4.5-d~imidazole (Compound 1) ~3 A mixture of 2,2'-pyridil (15.8 g, 74.4 mmol) and 2-guanidinobenzimidazole (19.5 g, 111.7 rnmol) in m~.th~nol (440 mL) was treated with a solution of sodium hydroxide (2.97 g, 74.4 mmol) in 74 mL and the resulting mixture was left st~n-ling at room te~ er~lulc for 4 days. The crystalline m~tçri~l was filtered and dried under vacuum to yield 21.1 g of the title compound as off-white crystals (72%). mp: 305-307 ~C (dec); lH NMR t300 MHz, d6-DMSO) ~ 11.5 (br s, NH, 2 H), 10.0 (br s, . . .

wo 97/44033 PCT/US97/08864 NH,2H),8.6(brs,NH,2H),8.38(d,J=4.2Hz,2H),7.55(t,J=7.8Hz,2H), 7.29(d,J=7.8Hz,2H),7.27-7.21 (m,4H),7.14(brs,2H),6.98(dd,J=5.8,3.2 Hz, 4 H); MS (ESI) m/z 527 [M + H]+; Anal. Calcd. for C28H22N12 2/3H20: C, 62.44; H, 4.37. N, 31.21; Pound: C, 62.72; H, 4.08; N, 30.86.
s Example 2 tion of 2.5-Bis~2-benzimidazolyliminol-3a,6a-diphenyl-1.233a.4.5.6.6a-octahydroimidazor4.5-dlimida_ole (Compound 2) \)_ HN
NH
~ H--~\ ~N~

A mixture of benzil (1.05 g, 5.0 mmol) and 2-guanidinobçn7imi~1~7ole (1.57 g, 9.0 mmol) in benzene (25 mL) was refluxed in pyridine (10 mL) for 1 h. After evaporating most of the pyridine under reduced pressure, the residue was treatedwith hot toluene and the resultin~ precipitate was filtered. The pl~ci~ le was then dissolved in 9: 1 water:acetic acid (30 mL); the solution was filtered and the fi}trate 15 was neutralized to pH 7 with phosphate buffer. A precipitate formed, that was then collected and triturated with water to afford the title compound (0.42 g, 16%).1H
NMR (300 MHz, d6-DMSO) o 11.5 (br s, NH, 2 H), 10.0 (br s, NH, 2 H), 8.6 (br s, NH, 2 H), 7.28-7.10 (m, 14 H), 6.97 (dd, J = 6.0, 3.0 Hz, 4 H); MS (ESI) n~z 525[M + H]+; Anal. Calcd. for C30H24Nlo . 1/2 CH3CO2H . 3/4H2O: C, 65.37; H, 4.88; N, 24.65. Found: C, 65.36; H, 4.79; N, 24.48.

Example 3 Preparation of 2.5-bisr2-ben7imitl~7olyliminol-3a.6a-bis(2-furyl)-1.2.33a.4.5.6~6a-bis(2-pyridyl)octahydroimidazor4.5-d~irnidazole Following the procedure of Example 1, 2,2'-furil (1.91 g, 10 mmol) and 2-gu~nifiinoben7imiAAr701e (2.10 g, 12.0 mmol) in methanol (60 mL) was treated with a solution of sodium hydroxide (400 mg, 10 mmol) in 10 mL of water. The black plc;cipit~te was filtered, dissolved 30 mL of 10% acetic acid and filtered. The 5 filtrate was neutralized with aqueous sodium hydroxide. The title compound pl~ipi~aLed as a grey solid (105 mg, 10% yield). mp: 222-225 ~C (dec); MS (ESI) m/z 505 [M + H]+.

Example 4 l~cp~uation of 2.5-bisr2-bel-7inA,il1~7.Qlyliminol-3a.6a-bis(3-metho~y~henyl)-1 .2.3.3a.4.5.6.6a-octahydroimicl~7.or4.5-dlimidazQle Following the procedure of Example 1, 3,3'-~limrAthoxybenzil (1.0 g, 3.69 mmol) and 2-guanidinobel-7;..1i~1 -7.~1e (540 mg, 3.08 mmol) in meth, nol (40 mL) was treated with a solution of sodium hydroxide (200 mg, 5 rnmol) in 5 rmL of water. The title compound was isolated as pale yellow crystals (655 mg, 73%
yield). mp: 276-278 ~C (dec);MS (ESI) m/z 585 [M + Hl+.

F.~mrle S

P~,p~lion of 2.5-bisr2-b~ -id~7,olylimino~-3a.6a-bis(4-~ ylp~.-yl)-1 .2.3.3a.4.5.6.6a-octahydroimidazor4.5-dlimid~7.ole Following the procedure of Example 1, 4,4'-dimethylbenzil (1.91 g, 12 mmol) and 2-guanidinoben~.i...id,A.7Ole (2.10 g, 10 mmol) in m,th~nol (60 mL) was treated with a solution sodium hydroxide (400 mg, 10 mmol) in 10 mL of water.
The orange solid was filtered, dissolved 40 mL of 10% acetic acid and filtered. The filtrate was then neutralized with aqueous sodium hydroxide to yield the title co-~ou~ld p-eeipi~ as a pale yellow solid (185 mg, 7~o yield). mp: 245-248 ~C
(dec); MS (ESI) m/z 553 [M + H]+.
Fx~ le 6 Plep~ation of 2.5-bisr2-be~.7i~ 7.olyliminol-3a.6a-bis(4-fluorophenyl)-1.2.3.3a.4.5.6.6a-octah~ydroimidazor4.5-dlimidazole Following the procedure of Example 1, 4,4'-difluorobenzil (1.0 g, 4.06 rnmol) and 2-g~l~ni~inobe-n~i...;~1~7,o'A (0.6 g, 3.4 mmol) in methanol (40 mL) was treated with a solution of sodium hydroxide (200 mg, 5 mmol) in 5 mL of water.

The ~itle compound was isolated as pale yellow crystals (750 mg, 79% yield). MS
(ESI) m/z 561 [M + H]+.

Example 7 ~ep~dlion of 2.5-bisr2-benzimida_olyliminol-3a.6a-bis(6-methyl-2-pvridyl)-I .2.3.3a.4.5.6.6a-octahydroimida_ol4.5-dlirnida_ole Following the procedure of Example 1, 6,6'-dimethyl-2,2'-pyridil (100 mg, 0.41 mmol) and 2-guanidinobe~7imi~i~7ol~ (60 mg, 0.35 mmol) in methanol (5 mL) was treated with a solution of sodium hydroxide (20 mg, 0.55 mmol) in 0.5 rnL of water. The title compound was isolated as pale yellow crystals (55 mg, 28%
yield). MS (ESI) m/z 555 [M + H]+.

Example 8 ion of 2.5-bisr2-be~ "id~7olylirr~nol-3a-(2-pyridyl)-6a-phenyl-1.2.3.3a.4.5.6.6a-octahydroimidazor4.5-d~imid~7ole Following the procedure of Example 1, 1-phenyl-2-(2-pyridinyl)eth~n~ione (800 mg, 3.79 rnrnol) and 2-guanidinoben7imiA~701e (560 mg, 3.15 mmol) in methanol (20 mL) was treated with a solution of sodium hydroxide (40 mg, 1.0 mmol) in 1 mL of water. The title cu~llpo-nd was isolated as pale yellow crystals (160 mg, 20% yield). lH NMR (300 MHz, d6-DMSO) ~
12.0 (br. s, NH, lH) 11.5 (br s, NH, 2 H~, 10.0 (br s, NH, 1 H), 8.6 (br s, NH, 1 H), 8.41 (m, 2 H), 7.55 (t, J= 6.9 Hz, 2 H), 7.30-7.10 (m, 8 H), 6.98 (dd, J= 6.0, 3.1 Hz, 4 H); MS (ESI) m/z 526 [M + H]+.

Exarnple 9 ~p~dlion of 2.5-bisr2-bel~7;~ 7olyliminol-3a-(4-pyridyl)-6a-phenyl-1 ~ ~.3a.4~5.6.6a-octal.~.l,uhllidazor4~5-d]imidazole Following the procedure of Example 1, 1-phenyl-2-(4-pyridinyl)eth~n~ione (500 mg, 2.42 rnmol) and 2-guanidinobel-7i---ifi~7(-1e (1.0 g, 6.0 mmol) in methanol (10 mL) was treated with a solution of sodium hydroxide (40 mg, 1.0 rnmol) in 1 mL of water. The title compound was isolated as pale yellow crystals (45 mg, 3% yield). lH NMR (300 MHz, d6-DMSO) ~ 11.5 ~br s, NH, 1 H), 11.0 (br.s, NH, lH), 10.0 (br s, NH, 1 H), 8.6 (br s, NH, 1 H), 8.4 (m, 2 H), 7.4-6.8 (m, l5H); MS (ESI) n~z 526 [M + H]+.

Example 10 Preparation of 2.5-bisr(5-methyl-2-ben7imid~7.olyl)imino]-3a.6a-bis(2-pyridyl)-1 23.3a.4.5.6.6a-octahydroimi~7,c)r4.5-dlimidazole Following the procedure of Example 1, 2,2'-pyridil (603 mg, 2.84 mmol) and 5-methyl-2-guanidinoben7imi~7ole (489 mg, 2.58 mmol) in methanol (17 mL) was treated with a solution of sodium hydroxide (113.6 mg, 2.84 mmol) in 2.8mL of water. The title co~ o~ d was isolated as a grey powder (450 mg, 63%
yield). mp: 290-291 ~C (dec); IH NMR (300 MHz, d6-DMSO) o 11.3 (br s, NH, 2 H), lO.O(brs,NH,2H),8.5(brs,NH,2H),8.32(d,J=4.2Hz,2H),7.54(t,J=

7.6Hz,2H),7.31 (d,J=7.6Hz,2H),7.16(d,J=7.8Hz,2H),7.16-7.09(m,2 H),7.09(s,2H),7.14(brs,2H),6.86(d,J=7.8,Hz,2H),2.34(s,6H);MS
(ESI) m/z 555 [M + Hl+
Example 11 ~paldlion of 2.5-bis~(5-nitro-2-ben7imitl~7.oly1)iminol-3a.6a-bis(2-pyridyl)-1.2.3.3a.4.5.6.6a-octahydroimida_or4.5-dlimida_ole Following the procedure of Example 1, 2,2'-pyridil (245 mg, 1.15 mrnol) and 5-nitro-2-guanidinoben7.imi~701e (254 mg, 1.15 mmol) in methanol (6.9 mL) was treated with a solution of sodium hydroxide (46 mg, 1.15 mmol) in 1.1 mL of water. The title compound was isolated as orange crystals (182 mg, 51%). mp-340-350 ~C (dec); MS (ESI) nz/z 617 IM + Hl+.
F.x~m~l?le 12 Preparation of 2.5-bisr(5-chloro-2-be~ 7.olyl)iminol-3a.6a-bis(2-pyridyl)-1.2.3.3a.4.5.6.6a-octahydroimida_o~4.5-dlimi-1~7c)le~
Following the procedure of F.Y~mpl~ 1, 2,2'-pyridil (352 mg, 1.66 mmol) and 5-methyl-2-guanidinobe~7imi~l~7Ole (350 mg, 1.66 mmol) in m~th~nol (10 mL) was treated with a solution of sodium hydroxide (66.4 mg, 1.66 mmol) in 2.5 mL of water. The title compound was isolated as a brownish solid (247 mg, 50%).
MS (ESI) m/z 596 lM + Hl+
Example 13 , .. , . . ~ ~ ... . .

CA 022~731 1998-11-20 Preparation of 2.5-bisr(4-methyl-2-thiazolyl)iminol-3a 6a-bis(2-pyridyl)-1.2.3.3a~4~5 ~6a-octahydroimidazor4~5-dlimidazole Following the procedure of Example 1, 2,2'-pyridil (1.06 g, 5.0 mmol) and 4-methylthia7Ol-2-ylgu~nitline hydrochloride (1.35 g, 7.0 mmol) in methanol (30 S mL) was treated with a solution of sodium hydroxide (450 mg, 11.25 m~nol) in 7.5 mL of water. The ~l~cipilate was filtered, dissolved 30 mL of 10% acetic acid and filtered; the filtrate was then neutralized with aqueous sodium hydroxide. The title compound precipitated as a tanned solid (265 mg, 16% yield). mp: >300 ~C (dec);
MS (ESI) nz/z 489 [M + H]+.
Example 14 Preparation of 2~5-bisr2-benzoxazolylimino~-3a~6a-bis(2-pyridyl)-1~23a.4.5.6.6a-octahydroimi~l~7.Qr4.5-dlimida_ole Following the procedure of FY~mple 1, 2,2'-pyridil (301 mg, 1.42 mmol) and 2-guanidinobenzoxazole (250 mg, 1.42 mmol) in methanol (12 mL) was treated with a solution of sodium hydroxide (56.7 mg, 1.42 mmol) in 2 mL of water. The title compound was isolated as a brownish solid (151 mg, 40%). 1H
NMR (300 MHz, d6-DMSO) ~ 10.28 (s, NH, 4 H), 8.36 (d, J = 4.6 Hz, 2 H), 7.57 (td, J= 7.8 1.8 Hz, 2 H), 7.47 (d, J= 7.8 Hz, 2 ),7.39 (d, J= 7.8 Hz, 2 H), 7.30 (d, J= 4.6 ~Iz, 2 H), 7.19 (dt, J = 7.7, 1.3 Hz, 2 H), 7.16-7.10 (m, 4 H); MS (ESI) m/z 529 [M + H]+-Example 15 ~palalion of 2.5-bisr(4-methyl-2-benLin~idazolyl)imino1-3a.6a-bist2-pyridyl)-1.2.2a.4.5.6.6a-octahydroimidazo~4.5-dlimida_ole Following the procedure of Example 1, 2,2'-pyridil (1.87 g, 8.83 mmol) and 4-methyl-2-g-~nillinQbe~7imi~7ole (2.0 g, 10.5 mmol) in m~th~nol (63 mL) was treated with a solution of sodium hydroxide (420 mg, 10.5 mmol) in 6 mL of water. The title compound was isolated as white crystals (1.9 g, 65%). MS (ESI) n2/z 555 [M + H]+.

Example 16 Preparation of 5-r2-b~n~.imiri~7olylitninol-2-~(4-methyl-2-be,lzi",idazolyl)imino~-3a.6a-bis(2-pyridyl)- 1 .2.2a.4.5,6~6a-octahydroimidazor4.5-dlimidazole bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (135 mg, 0.636 mrnol), 2-guanidinobenzimidazole (92.8 mg, 0.530 mmol) and 4-methyl-2-guanidinobP.n7.imi~7.01e (100 mg, 0.530 mmol) in m~th~rlol (3 mL) was treated with a solution of sodium hydroxide (38 mg, 0.95 mmol) in 0.5 mL of water and the resulting mixture was left st:~n-ling at room te,ll~elaLu~c for 2 days. The crystalline material was filtered and purified by reversed phase ~c~a~ e HPLC (Rainin Dynamax, S IlM C18 column: 21.4 mm x 25 cm, elution with gradient ncetonitrile-water cont~ining 0.1 % trifluoroacetic acid) to yield the title compound as a white powder (100 mg, 25%). MS (ESI) m/z 541 ~M + Hl+-Example 17 F~p~alion of 5-(2-ben7i~ 7.olylimino)-2-r(5-methyl-2-benzimidazolyl)iminol-3a.6a-bis(2-pyridyl)- 1 ~2.3.3a~4.S.6.6a-octahydroimidazor4.5-dlimidazole bis(trifluoroacet~t~) salt Following the procedure of Example 16, except subsliluling 5-methyl-2-guanidinoben7;.lli-1~70le for 4-methyl-2-gu~ni~1inobe~ 17.i ~-.id~7O1e, the title compound was prtpaled (88 mg, l l~ H NMR (300 MHz, d6-DMSO) o 13.0 (brs,NH,4H),9.8(brs,NH,4H),8.39(d,J=4.3Hz,2H),7.64(td,J=7.8, 1.7 Hz, 2 H), 7.57 (d, J = 7.8 Hz, 2 H), 7.49-7.46 (m, 2 H), 7.38-7.33 (m, 3 H), 7.27 (s, 1 H), 7.21-7.13 (m, 3 H), 2.44 (s, 3 H); MS (ESI) m/z 541 EM + Hl+.
Example 18 ationof2.5-bisr(l-methyl-2-bc,lzil,lidazolyl)imino]-3a.6a-bis~2-pyridyl)-1.2.3.3a.4.5.6.6a-octahyd,~ idazo~4.5-dlimid~ole bisCtriflllnro~cetat~) salt Following the procedure of F.l~mple 1, 2,2'-pyridil (187 mg, 0.88 mmol) and l-methyl-2-guanidinoben7i~ 7-)le (0.20 g, 1.57 mmol) in methanol (6 mL) was treated with a solution of sodium hydroxide (60 mg, 1.5 mmol) in 0.6 rnL of water. The crystalline material was filtered and purified by reversed phase dli~e HPLC (Rainin Dynamax, 5 ~lM C18 column: 21.4 mm x 25 cm, elution with gradient acetonitrile-water cont~inin~ 0.1% trifluoroacetic acid) to yield the title compound as a white powder (200 mg, 48~?o). lH NMR (300 MHz, d6-DMSO) o 9.9 (br s, NH, 4 H), 8.41 (d, J = 4.5 Hz, 2 H), 7.68-7.62 (m, 4 H), 7.54 (d, J = 7.7 Hz, 2 H), 7.55-7.50 (m, 2 H), 7.39-7.36 (m, 4 H), 7.20 (ddd, J =
7.3, 4.9, 1.0 Hz, 2 H), 3.78 (s, 6 H); MS (ESI) n~z 555 [M + Hl+.

Example 19 Prcparation of 5-(2-ben7imidazolylimino)-2-rr5-fluoro-2-benzimidazoly3a.6a-bis(2-pyridyl)- l .2.3.3a,4.5.6.6a-octahydroimidazor4.5-dlimidazole bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (345 mg, 1.6 mmol), 2-gu~ni~linobe~ 7.ole (181 mg, 1.036 mmol) and 5-fluoro-2-guanidinoben7.imi~7O1e (250 mg, 1.036 mmol) in mçth~nol (3 mL) was treated with a solution of sodium hydroxide (60 mg, 1.5 mmol) in 0.5 mL of water and the resulting mixture was left st~n-ling atroom te~ e.~lulc for 2 days. The crystalline m~teri~l was filtered and purified by reversed phase ~ ~ati~e HPLC (Rainin Dynamax, S llM C18 column: 21.4 mm x 25 cm, elution with gradient acclollillile-water co~";.ini--~ 0.1% trifluoroacetic acid) to yield the title compound as a white powder (92 mg, 12%); lH NMR (300 MHz, d6-DMSO) o 13.2 (br s, NH, 2 H), 9.8 (br s, NH, 4 H), 8.40 (d, J = 4.2 Hz, 2 H), 7.64 (td, J = 7.6, 1.2 Hz, 2 H), 7.55 (d, J = 7.5 Hz, 2 H), 7.49-7.43 (m, 3 H), 7.34-7.30 (m, 3 H), 7.19 (dd, J = 7.0, 5.0 Hz, 2 H); MS (ESI) ~z/z 545 [M + H]+.
Exarnple 20 ~ep~dlion of 2~5-bisr(5-fluoro-2-bel,~ill~idazolyl)iminol-3a~6a-bis(2-pyridyl)-1~233a~4~5~6~6a-octahydroimidazo~4.5-dlirnicl~7.nle bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (262 mg, 1.2 mmol) and 5-fluoro-2-gu~ni~linc be~ 7Ole (200 mg, 1.04 mmol) in mP.th~nol (3 mL) was treated with a solution of sodium hydroxide (60 mg, 1.5 mmol) in 0.6 mL of water. The crystalline m~tPri~l was filtered and purified by reversed phase plc~ali~/e HPLC(Rainin Dynamas, 5 ~lM C18 column: 21.4 mm x 25 cm, elution with gradient acelollillile-water cont:3ining 0.1% tlifluoroacetic acid) to yield the title compound as a white powder (147 mg, 37%). 1H NMR (300 MHz, d6-DMSO) ~ 13.0 (br s, NH,2H),9.9(brs,NH,4H),8.39(d,J=4.6Hz,2H),7.64(td,J=7.7 1.6Hz,2 H), 7.52(d,J=7.8Hz,2),7.45-7.41 (m,2H),7.32-7.28(m,2H),7.21-7.16(m, 2 H), 7.15-7.11 (m, 2 H); MS (ESI) m/z 563 [M + H]+.
Example 21 . ~ . . .

Wo 97/44033 PCT/US97/08864 Preparation of 5-(2-bçn7.imi~7olylimino)-2-r(l-methyl-2-ben7.imid~7olyl)iminol-3a.6a-bis(2-pyridyl)- 1.2.3.3a.4.5.6~6a-octahydroirnidazor4.5-dlimidazole bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (265 mg, 1.25 mmol). 2-guanidinobçn7imitl~7.01e (231 mg, 1.32 mmol) and 1-methyl-2-guanidinobenzimidazole (350 mg, 1.59 mrnol) in methanol (3 mL) was treated with a solution of sodium hydroxide (95 mg, 2.4 mmol) in 0.5 mL of water and the resulting mixture was left ct~nfling atroom lelllp.,lature for 2 days. The crystalline material was filtered and purified by reversed phase pl~;p~àlive HPLC (Rainin Dynamax, 5 ~M C18 column: 21.4 mm x 25 cm, elution with gradient acetonitrile-water cont~ining 0.1% trifluoroacetic acid) to yield the title compound as a white powder (125 mg, 14%). lH NMR (300 MHz, d6-DMSO) ~ 13.2 (br s, NH, 2 H), 9.9 (br s, NH, 4 H), 8.40 (d, J = 4.3 Hz, 2 H), 7.68-7.62 (m, 3 H), 7.57 (d, J = 7.8 Hz, 2 H), 7.50-7.47 (m, 3 H), 7.38-7.30 (m, 4 H), 7.20 (ddd, J = 7.3, 5.3, 1.0 Hz, 2 H),, 3.78 (s, 3 H); MS (ESI) m/z 541 [M +
Hl+.

Fx7mRle 22 Preparation of 5-~2-benzimidazolyliminol-2-r(4-methyl-2-thiazolyl)iminol-3a.6a-bis(2-pyridyl)-1.2~2a.4.5.6 6a-octahydroimudazo~14 5-dlimidazole bis(trifluoroacetate) salt Following the procedure of F.lr~mple 19 except substituti~g 4-1lylthiazol-2-yl~ ni~line for 5-fluoro-2-guanidinobenzimidazole, the title colllpoulld was prepared (63 mg, 7%). MS (ESI) n~z 510 [M + Hl+.
Example 23 ~p~tion of2~5-bisr(5-bromo-4-methyl-2-thiazolyl)iminol-3a.6a-bis(2-wridyl)-1 ~ ~.3a.4.5.6a-octahydroimidazor4.5-dlimid~7.ole Following the procedure of Example 1, 2,2'-pyridil (215 mg, 1.016 mmol) and 5-bromo-4-methyl-2-guanidinoben7imi~7O1e (200 mg, 0.847 mmol) in m~h~nol (3 mL) was treated with a solution of sodium hydroxide (51 mg, 1.27 mrnol) in 0.5 mL of water. The title compound was isolated as white crystals (50mg, 18%). MS (ESI) m/z 647 [M + H3+.
Fx~mrle 24 ion of 5-r2-ben7imid ~zolyliminol-2-r(5-chloro-2-benzimida_olyl)iminol-3a.6a-bis(2-pyridyl)- 1.2.2a.4.5.6.6a-octahydroimidazor4.5-dlimida_ole bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (242 mg, 1.14 mmol), 2-guanidinobçn7imirlA70l~
(166 mg, 0.95 mmol) and 5-chloro-2-guanidinobe.. ii---i~A7.01e (200 mg, 0.95 mmol) in mPthAnol (3 mL) was treated with a solution of sodium hydroxide (57 mg, 1.42 mmol) in 0.5 rnL of water and the resulting mixture was left stAn-ling at room le~ dtu~t for 2 days. The crystalline material was filtered and purified byreversed phase ~lcp~alive HPLC (Rainin Dynamax, 5 ~M C18 column: 21.4 mm x 25 cm, elution with gradient acetonitrile-water contAining 0.1% trifluoroacetic acid) to yield the title eolllpoulld as a white powder (173 mg, 23%). MS (ESI) m/z 562 [M + H}+.

Example 25 Preparation of 2.5-bisr(5.6-dimethyl-2-ben~ A7c 1yl)iminol-3a.6a-bis(2-pyridyl)-1.2.2a.4.5.6.6a-octahydroimidazo~4.5-dlimida_Qle bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (254 mg, 1.2 mmol) and 5,6-dillletllyl-2-guanidinoben7.imi(1~701e (203 mg, 1.0 mmol) in methanol (3 mL) was treated with a solution of sodium hydroxide (60 mg, 1.5 mmol) in 0.6 mL of water. The crystalline mAtPriAl was filtered and purified by reversed phase ~pa.aLive HPLC
(Rainin Dynamax, 5 ,uM C18 column: 21.4 mrn x 25 cm, elution with gradient ~to~ -water co~ g 0.1 % trifluoroacetic acid) to yield the title compound as a white powder (170 mg, 42%). MS (ESI) ~z 583 [M + Hl+.

Example 26 p~aLion of 5-r2-benzimida_olyliminol-2-r(5.6-dimethyl-2-b.on7imi~l~7olyl)iminol-3a.6a-bis(2-pyridyl)-1.2.2a.4.5.6.6a-octA~wdroirnida_or4.5-dlimidazole bis(trifluoroacelale) salt A mixture of 2,2'-pyridil (254 mg, 1.2 mrnol), 2-guanidinobç~,.i-.-iclA7.0le (175 mg, 1.0 mmol) and 5,6-dimethyl-2-guanidinoben7.imi~1~7O1e (203 mg, 1.0 m~nol) in m-o.thAnol (3 ml) was treated with a solution of sodium hydroxide (60 mg, 1.5 m~nol) in 0.5 mL of water and the resl-lting mixture was left ~tAn-ling at room te~ elà~ulG for 2 days. The crystalline material was filtered and purified by reversed phase pl~;p~alive HPLC (Rainin Dynamax, 5 ~lM C18 column: 21.4 mm x 25 cm, elution with gradient aretonitrile-water ContAinin~ 0.1% trifluoroacetic acid) to yield the title compound as a white powder (96 mg, 12%). IH NMR (300 MHz,d6-DMSO)o 13.1 (brs,NH,2H), 12.9(brs,NH,2H),9.9(brs,NH,2H), 9.8(brs,NH,2H),8.39(d,J=4.3Hz,2H),7.64(td,J=7.9,1.5Hz,2H), 7.58 ~d,J=7.9Hz,2H),7.48(dd,J=6.0,3.2Hz,2H),7.34-7.31 (m,2H),7.25 (s,2 H), 7.19 (dd, J= 7.0, 5.0 Hz, 2 H), 2.33 (s, 6 H); MS (ESI) n~z 555 [M + H~+.

Example 27 ~l~pa,dlion of 2.5-bisr(5-carboxy-2-be~7imi~l~zQlyl)iminol-3a6a-bis(2-pyridyl)-1~2~2a~4~5~6~6a-octahydroimidazor4~5-dlimid~ole bis(trifluoroacetate) salt Following the procedure of P.Y~mple 25 except substituting 4-carboxy-2-gu~,ni-linoben~.;...id~701e for 5,6-dill~clhyl-2-gl~~ni-linobeh~ lA7ole~ the title compound was prepared after pl-riflc~tion by reversed phase plcp~dLi.le HPLC (42mg, 10%). MS (ESI) m/z 615 [M + Hl+.

Exarnple 28 P~ tion of 5-r2-ben7in~ 7Qlyliminol-2-r2-benzoxazolyliminol-3a.6a-bis(2-pyridyl)-1.~ ~a~4~5.6~6a-octahydloi,.lidazor4.5-dlimidazole bis(trifluoroacetate) salt A l"i~Lu~c of 2,2'-pyridil (143 mg, 0.68 mmol), 2-g~1~ni~1ino~el-7.;.~ 7.01~.
(100 mg, 0.56 mmol) and 2-guAni(1inobenzoxazole (100 mg, 0.56 mmol) in mPth~nol (3 mL) was treated with a solution of sodium hydroxide (34 mg, 0.85 mmol) in 0.5 mL of water and the reslllting mixture was left stAn~ling at room tclll~el~llJre for 2 days. The crystalline m~t~.ri~l was filtered and purified by reversed phase pl~a~alive HPLC (Rainin Dynamax, 5 ~IM C18 column: 21.4 mm x 25 cm, elution with gradient ~cetonitril~-water col~lAil~ine 0.1% trifluoroacetic acid) to yield the title compound as a white powder (42 mg, 10%). MS (ESI) m/z 528 [M + Hl+.

Example 29 P~ J~alionof 2.5-bisr2-benzothiazolyliminol-3a~6a-bis(2-pyridyl)-1.2.3a.4.5.6.6a-octahydroimidazo~4.5-dli~ A7ole.
A mixture of 2,2'-pyridil (212 mg, 1.0 mmol) and 2-guanidinobenzothiazole (403 mg, 2.2 rnmol) in methanol (10 mL) was treated with a solution of sodium hydroxide (30 mg, 0.75 mmol) in 0.3 rnL of water. The titlecompound was obtained as crystalline powder (407 mg, 73%). IH NMR (300 MHz, d6-DMSO) o 10.3 (br s, NH, 2 H), 8.38 (d, J = 4.0 Hz, 2 H), 7.94 (s, J = 7.8, 2H),7.64(m,4H),7.41 (m,4H),7.30(t,J=7.5Hz,2H), 7.18(dd,J=7.5,4.9 5 Hz, 2 H); MS (ESI) m/z 561 [M + H~+.

Exarnple 30 Ple~ation of 5-~2-ben~ idazolyliminol-2-r2-benzothiazolyliminol-3a.6a-bis(2-pyridyl)-1.2.3a.4.5.6.6a-octahydroirnidazor4.5-dlimi~1~7ole bis(trifluoroacetate) salt A mixture of 2,2'-pyridil (255 mg, 1.2 mmol), 2-gu~ni-linobenzimida-ole (175.2 mg, 1.0 mmol) and 2-gl-~ni~inobenzothia_ole (192 mg, 1.0 mmol) in m.o.th~nol (10 mL) was treated with a solution of sodium hydroxide (40 mg, 1.0 mmol~ in 0.4 mL of water and the res~lting mixture was left st~n-lin~ at room ~tlllyeldlule for 2 days. The crystalline m~teri:~l was filtered and purified byreversed phase pl~a~ive HPLC (Rainin Dynamax, 5 IlM C18 column: 21.4 mm x 25 cm, elution with gradient acetonitrile-water cont~ining 0.1% trifluoroacetic acid) to yield the title compound as a white powder (30 mg, 4%) lH NMR (300 MHz, d6-DMSO) o 13.0 (br s, NH, 2 H), 9.8 (br s, NH, 2 H), 8.40 (d, J = 4.0 Hz, 2H),7.83(d,J=7.7,H),7.55(t,J=7.8Hz,2H),7.54(d,J=7.7Hz, 1 H),7.50-7.38 (m, 4 H), 7.38-7.25 (m, 3 H), 7.23-7.15 (m, H); MS (ESI) m/z 544 [M+ H]+.

Example 31 P~ ~alion of 5-~(2-ben7i",i-1~zolylimino)l-2-~(5-carboxy-2-7.i 11 ,i(1~7olyl)iminol-3a.6a-bis(2-pyridyl)- 1.2.3.3a.4.5.6~6a-octall~dloi ~ ";(1~7or4.5-dlimidazole bis(trifluoro~r-et~te) salt A mixture of 2,2'-pyridil (508 mg, 2.4 rnmol), 2-guanidinobe~-~il";r1~7ole (350 mg, 2.0 mmol) and 5-carboxy-2-guanidinoben~ idazole (428 mg, 2.0 mmol) in methanol (6 mL) was treated with a solution of sodium hydroxide (120 mg, 3.0 mmol) in 0.8 mL of water and the res--lting mixture was left st~ntlin~ at room tel,lpeld~ule for 2 days. A total of 490 mg of a crystalline mz-teri~l was obtained, a portion of which (250 mg) was purified by reversed phase ~ ~d~ive HPLC
(Rainin Dynamax, S IlM C18 column: 21.4 mm x 25 cm, elution with gradient acetonitrile-water cont~ining 0.1% trifluoroacetic acid) to yield the title culllpùund as a white powder (50 mg, 6%).MS (ESI) m/z 571 [M + H]+.

~. .

W O 97/44033 PCTrUS97/08864 Example 32 ~l~p~alion of 2.5-bisr(5-iodo-2-benzimidazolyl)iminol-3a.6a-bis(2-pyridyl)-1.2.3.3a.4.5.6.6a-octahydroimidazor4.5-dlimi(1~7ole bis(trifluoroacetate) salt and 5-~2-benzimidazolyliminol-2-~(5-iodo-2-benzimidazolyl~iminol-3a.6a-bis(2-pyridyl)- 1.2.2a~4.5.6.6a-octahydroimidazor4.5-dlimidazole bis(trifluoroacetate) salt A solution of 2,5-bis[2-~n7imi-1~7olylimino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imid~7.ole (2.0 g, 3.8 mmol) in a mixture of 35 mL of N,N-dimethyl formamide, 3.5 mL of acetic acid and 3.5 mL was treatedwith N-iodosuccinimide (1.71 g, 7.6 mmol) and the resulting mixture was stirred for 3 d at room t~ y~l~tur~. The solvent was evaporated and the residue partitioned between ethyl acetate and water; the aqueous layer was neutralized with sodium bicarbonate solution and e~traeled with ethyl acetate twice. The combined organic extracts were washed with water, brine and dried over anhydrous sodium sulfate.
The solvent was evaporated under reduced pressure to yield a solid material (3.2 g);
a portion of it (1.0 g) was purified by reversed phase pltp~dli~re HPLC (Rainin Dynamax, 5 ,uM C18 column: 21.4 rnm x 25 cm, elution with gradient acetonitrile-water Cont~ining 0.1% trifluoroacetic acid) to yield the title col~oLInds as white powders. 5-iodo derivative (12.5 mg, 1%): lH NMR (300 MHz, d6-DMSO) ~ 13.0 (brs,NH,2H),lO.O(brs,NH,2H), 8.39(d,J=4.7Hz,2H),7.64(t,J=5.9Hz,2 H), 7.40-7.60 (m, 4 H), 7.32 (m, 2 H), 6.98 (dd, J = 4.6, 3.2 Hz, 2 H); MS (ESI) m/z 653 [M + Hl+. 5,5'-diiodo derivative (62.5 mg, 12%): IH NMR (300 MHz, d6-DMSO) ~ 9.9 (br s, NH, 2 H), 8.40 (d, J = 4.5 Hz, 2 H), 7.74 (s, 2 H),7.62 (t, J =
5.9, 1.7 Hz, 2 H), 7.45-7.56 (m, 4 H), 7.28 (d, 2 H), 7.19 (m, 3 H); MS (ESI) m/z 779 [M + Hl+.

Example 33 Preparation of 2.5-bisr2-benzimidazoly~iminol-3a~6a-bis(4-bromophenyl)-1 .2.3.3a.4.5.6.6a-octahydroimidazor4.5-dlimidazole CA 022~731 1998-11-20 Following the procedure of Example 1, 4,4'-dibromobenzil (4.42 g, 12 mmol) and 2-guanidinoben7imi(l:l701e (1.75 g, 10 mmol) in methanol (100 mL) was treated with a solution of sodium hydroxide (400 mg, 10 mrnol) in 10 mL of water. The title compound was isolated as pale yellow crystals (800 mg, 24%
S yield). mp: 274-278 ~C (dec); MS (ESI) m/z 683 [M + H]+.

Example 34 Preparation of 2,5-bisr2-benzimidazolyliminol-3a.6a-bis(4-trifluoromethylphenyl)-1~2.3.3a~4.5.6.6a-octahydroimidazor4.5-dlimidazole Following the procedure of F.~mple 1, 4,4'-trifluoromethylbenzil (0.253 g, 0.73 mmol) and 2-guanidinobenzirnidazole (0.154 g, 0.88 mmol) in methanol (10 mL) was treated with a solution of sodium hydroxide (35.2 mg, 0.88 mmol) in 1 mL of water. The title compound was isolated as pale yellow crystals (180 mg, 15 62% yield). MS (ESI) m/z 661 [M + H]+.

Example 35 Preparation of 2.5-bisr2-ben~7imi-1~7.olyliminol-3a-(3-chlorophenyl)-6a-phenyl-1.2.3.3a~4.5.6.6a-octahydroirnidazor4.5-dlimidazole Following the procedure of Example 1, 1-(3-chlorophenyl)-2-phenylethanedione (0.25 g, 1.0 mmol) and 2-guanidinobenzimidazole (0.215 g, 1.2 mmol) in m~h~nol (12 mL) was treated with a solution of sodium hydroxide (48 mg, 1.2 mmol) in 1.2 mL of water. The title compound was i~ol:~ted as pale 25 yellow crystals (130 mg, 39% yield). MS (ESI) n~z 559 [M + H]+.

Example 36 Plepa~alion of 2.5-bisr2-be~7imid~7olyliminol-3a-(2-chlorophenyl)-6a-phenyl-1.2.3.3a.4.5.6.6a-octahydroimidazor4~5-dlimidazole Following the procedure of Example 1, 1-(2-chlorophenyl)-2-phenyleth~ne-liQn.o (0.25 g, 1.0 mmol) and 2-guanidinobenzirnidazole (0.215 g, 1.2 mmol) in methanol(12 mL) was treated with a solution of sodium hydroxide (48 mg, 1.2 mmol) in 1.2mL of water. The title compound was isolated as pale yellow crystals (130 mg, 35 39% yield). MS (ESI) n2/z 559 [M + H]+.

Example 37 Preparation of 5-(2-ben7.imiri~zQlylimino)-2-[(5-methyl-2-ben7imi(~7olyl)iminol-3a-(2-pyridyl)-6a-phenyl- 1 ~2.3.3a.4.5.6.6a-octahydroirnidazor4.5-dlimidazole bis(trifluoroacetate) salt A rnixture of 2-phenyl-1-(2-pyridyl)ethanedione (253 mg, 1.2 mmol), 2-guanidinoben7imi~7.01e (175 mg, 1.0 mmol) and 5-methyl-2-guanidinobçn7imiA~7Ole (189 mg, 1.0 rnrnol) in methanol (6 mL) was treated with a solution of sodium hydroxide (40 mg, 0.95 rnmol) in 1.0 mL of water and the res~llting mixture was left st~nrling at room telllpel~lurG for 2 days. The crystalline material was filtered and purified by reversed phase plGpa~ e HPLC (Rainin Dynarnax, 5 mM C18 colurnn: 21.4 mm x 25 cm, elution with gradient ~cct- nitrile-water cont~ining 0.1% trifluoroacetic acid) to yield the title colll~oulld as a white powder (20 mg, 5%). MS (ESI) nz/z 540 [M + H]+.

F~ramrle 38 ~alion of 2.5-bis~(l-methyl-2-b~.~zilllida_olyl)iminol-3a.6a-d~henyl-1.2.3.3a.4.5.6.6a-octahydromidazol4.5-dlirnidazole A solution of benzil (210 mg, 1.0 mmol) and 1-methyl-2-guanidinoben7i.... irl~7,ole (207 g, 1.1 mmol) in ethanol (5 mL) was treated with sodium hydroxide (40 mg, 1.0 mmol) in 0.8 mL of water and the resulting mixture was refluxed for 2 hours. The title compound yiGcipilated and was isolated as a light yellow powder (125 mg, 41% yield). MS (ESI) m/z 553 [M + H]+.

Example 39 udlion of 2.5-bisr(1-methyl-2-bellzill~idazolyl)iminol-3a.6a-bis(4-ln~;lllylphenyl)- 1 .2.3.3a.4.5.6.6a-octahydromidazor4,5-dlimida_ole A solu~ion of 4,4'-dimethylbenzil (238 mg, 1.0 mmol) and 1-methyl-2-guanidinobel.7.i.. ,i~l~7Ole (207 g, 1.1 mmol) in ethanol (5 mL) was treated with sodium hydroxide (40 mg, 1.0 mmol) in 0.8 mL of water and the resul~ing mixture was refluxed for 2 hours. The title compound precipitated and was isolated as a yellow powder (50 mg, 16% yield). MS (ESI) m/z 581 [M + H]+.

Example 40 Plc,~dlion of 2.5-bisr(l-methyl-2-benzimida_olyl)irninol-3a.6a-bis(4-bromophenyl)-1.2.3.3a.4.5.6 6a-octahydrornidazor4.5-dlimida_ole A solution of 4,4'-dibromobenzil (368 mg, 1.0 rnrnol) and 1-methyl-2-guanidinobenzirnidazole (207 g, 1.1 mrnol) in ethanol (5 mL) was treated with sodium hydroxide (40 mg, 1.0 rnmol) in 0.8 mL of water and the reslllting rnixture was refluxed for 2 hours. The title compound pr~ci~ilated and was isolated as a pale yellow powder (320 mg, 82% yield). MS (ESI) m/z 71 1 [M + H]+.

Exarnple 41 - Capsule Composition An oral dosage form for ~mini~tering a presently invented agonist of the G-CSF lGceplor is produced by filing a standard two piece hard gelatin capsule with the ingredients in the ~lopG,Lions shown in Table I, below.
Table I
INGE~F.nIENTS AMOUNTS
2,5-Bis[2-bPn7imi~l~7.olylimino]-3a,6a-bis(2-pyridyl)- 25 mg 1 ,2,3,3a,4,5,6,6a-octahydroirnidazo[4,5-d]imi(1~7.ole (C~ p(.~ d 1) T -~rtose 55 mg Talc 16 mg Magnesium Stearate 4 mg Example 42 - Injectable Parenteral Cornposition An injectable forrn for ~mini~t~.ring a presently invented agonist of the G-CSF receptor is produced by stirring 1.5% by weight of 2,5-Bis[2-ben7imit1~7.olylimino]-3a,6a-diphenyl- 1 ,2,3,3a,4,5,6,6a-octahydrl-imi~7.o[4,5-d]irruda_ole (Compound 2) in 10% by volume propylene glycol in water.

Ex~m~le 43 - Tablet Col"posilion The sucrose, c~lcium sulfate dihydrate and a presently invented agonist of the G-CSF receptor, as shown in Table II below, are mixed and gr~nnl~t~d in the W O 97/44033 PCT~US97/08864 ~o~ollions shown with a 10% gelatin solution. The wet granules are screened, dried, mixed with the starch, talc and stearic acid, screened and compressed into a tablet.
Table Il s INGREDENTS AMOUNTS
2,5-Bis~2-ben7.imi-i~7,olylimino]-3a,6a-bis(2-pyridyl)- 20 mg 1 ,2,3,3a,4,5,6,6a-octahydroimid~7.o[4,5-d]imida_ole (Compound 1) calcium sulfate dihydrate 30 mg sucrose 4 mg starch 2 mg talc 1 mg stearic acid 0.5 mg While the plefell~,d embo-lim~n~ of the invention are illustrated by the above, it is to be understood that the invention is not limited to the precise instructions herein disclosed and that the right to all modifications coming within the 10 scope of the following claims is reserved.

... . .. ~ .

Claims (17)

What is claimed is:

1. A compound of the Formula (1):

wherein R1, R2, R3 and R4 are independently aryl, where aryl is cyclic or polycyclic aromatic C3-C12, optionally containing one or more heteroatoms, provided that when C is 3 the aromatic ring contains atleast two heteroatoms, and when C is 4 the aromatic ring contains at least one heteroatom, and optionally substituted with one or more substituents selected from the group consisting of: C(O)NR6R7 and NR6R7, aryloxy, cycloalkyl, substituted cycloalkyl, alkyl, C6-C12aryl, alkoxy, acyloxy, substituted C6-C12aryl, amino, N-acylamino, nitro, cyano, halogen, hydroxy, -C(O)OR6, -S(O)n R5, protected -OH
and alkyl substituted with one or more substituted selected from the group consisting of: alkoxy, acyloxy, C6-C12aryl, substituted C6-C12aryl, amino, N-acylamino, oxo, hydroxy, cycloalkyl, substituted cycloalkyl, -C(O)OR6, -S(O)n R7, aryloxy, nitro, cyano, halogen and protected -OH, where R6 is hydrogen or alkyl, n is 0-2, R7 is hydrogen or alkyl and R5 is hydrogen, cycloalkyl, C6-C12aryl, substituted cycloalkyl, substituted C6-C12aryl, alkyl or alkyl substituted with one or more substituents selected from the group consisting of: alkoxy, acyloxy, aryloxy, amino, N-acylamino, oxo, hydroxy, -C(O)OR6, -S(O)n R7, nitro, cyano, cycloalkyl, substituted cycloalkyl, halogen, C6-C12aryl, substituted C6-C12aryl and protected -OH, where R6 is hydrogen or alkyl, n is 0-2 and R7 is hydrogen or alkyl; and pharmaceutically acceptable salts, hydrates, solvates and esters thereof.

2. A compound of claim 1 in which R1 and R2 are independently phenyl, furyl, thienyl or pyridyl all of which are unsubstituted or substituted with a substituent selected from the group consisting of: halogen, C1-5alkyl, C3-6cycloalkyl and -O-C1-3alkyl;
R3 and R4 are independently pyridyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, thiazolyl or imidazolyl all of which are unsubstituted or substituted with one or two substitutents selected from the group concicting of: halogen, amino, cyano, N-acylamino, C1-3 alkyl, -O-C1-3 alkyl, nitro, -CO2H and CF3; and pharmaceutically acceptable salts, hydrates, solvates and esters thereof.

3. The compound of claim 1 selected from:
2,5-bis[2-benzimidazolylimino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a-(2-pyridyl)-6a-phenyl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 5-[2-benzimidazolylimino]-2-[(4-methyl-2-ben7imidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 5-(2-benzimidazolylimino)-2-[(5-methyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(1-methyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydromidazo[4,5-d]imidazole bis(trifluoroacetate) salt, and 5-[2-benzimidazolylimino]-2-[(5,6-dimethyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, or a pharmaceutically acceptable salt, hydrate or solvate thereof.

4. The compound of claim 1 selected from:

2,5-bis[2-benzimidazolylimino]-3a,6a-diphenyl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-bezimidazolylimino]-3a,6a-bis(2-furyl)-1,2,3,3a,4,5,6,6a-bis(2-pyridyl)octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-bezimidazolylimino]-3a,6a-bis(3-methoxyphenyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a,6a-bis(4-methylphenyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a,6a-bis(4-fluorophenyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a,6a-bis(6-methyl-2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a-(4-pyridyl)-6a-phenyl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[(5-methyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[(5-nitro-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[(5-chloro-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[(4-methyl-2-thiazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzoxazolylimino]-3a,6a-bis(2-pyridyl)-1,2,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[(4-methyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 5-(2-benzimidazolylimino)-2-[(5-fluoro-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(5-fluoro-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoluacetate) salt, 5-(2-benzimidazolylimino)-2-[(1-methyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 5-[2-benzimidazolylimino]-2-[(4-methyl-2-thiazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(5-bromo-4-methyl-2-thiazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6a-octahydroimiadazo[4,5-d]imidazole, 5-[2-bezimidazolylimino]-2-[(5-chloro-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(5,6-dimethyl-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(5-carboxy-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 5-[2-benzimidazolylimino]-2-[2-benzoxazolylimino]-3a,6a-bis(2-pyridyl)-1,2,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidaole bis(trifluoroacetate) salt, 2,5-bis[2-benzothiazolylimino]-3a,6a-bis(2-pyridyl)-1,2,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 5-[2-benzimidazolylimino]-2-[2-benzothiazolylimino]-3a,6a-bis(2-pyridyl)-1,2,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt 5-[(2-benzimidazolylimino)]-2-[(5-carboxy-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(5-iodo-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, and 5-[2-benzimidazolyliminio-2-[(5-iodo-2-benzimidazolyl)imino]-3a,6a-bis(2-pyridyl)-1,2,2a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, or a pharmaceutically acceptable salt, hydrate or solvate thereof.

5. A pharmaceutical composition comprising a pharmaceutically acceptable carrier and a compound of Claim 1.

6. A pharmaceutical composition for use in enhancing leukocyte production which comprises a compound of Claim 1 and a pharmaceutically acceptable carrier.

7. A pharmaceutical composition for use in treating bacterial infections which comprises a compound of Claim 1 and a pharmaceutically acceptable carrier.

8. A pharmaceutical composition for use in treating fungal infections which comprises a compound of Claim 1 and a pharmaceutically acceptable carrier.

9. A method of enhancing leukocyte production in a subject which comprises administering to the subject a therapeutically effective amount of a compound of Claim 1.

10. A method of treating neutropenia in a subject which comprises administering to the subject a therapeutically effective amount of a compound ofClaim 1.

11. A process for the preparation of a compound of Formula (I) wherein R1, R2, R3 and R4 are as described in claim 1; which comprises:
reacting a 1,2-diketone of Formula (2) wherein R1 and R2 are as described in Claim 1 with one or more guanidine derivatives of Formula (3) wherein R5 is R3 or R4, such that when more than one guanidine derivative of Formula (3) is utilized R5 does not have to be the same substituent, in a refluxing solvent with azeotropic removal of water, and thereafter optionally forming a pharmaceutically acceptable salt, hydrate orsolvate thereof.

12. A process for the preparation of a compound of Formula (I) wherein R1, R2, R3 and R4 are as described in claim 1; which comprises:
reacting a 1,2-diketone of Formula (2) wherein R1 and R2 are as described in Claim 1 with one or more guanidine derivatives of Formula (3) wherein R5 is R3 or R4, such that when more than one guanidine derivative of Formula (3) is utilized R5 does not have to be the same substituent, the presence of a base catalyst in a suitable protic solvent, and thereafter optionally forming a pharmaceutically acceptable salt, hydrate orsolvate thereof.

13. The compound of claim 1 selected from:

2,5-bis[2-benzimidazolylimino]-3a,6a-bis(4-bromophenyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a,6a-bis(4-trifluoromethylphenyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a-(3-chlorophenyl)-6a-phenyl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 2,5-bis[2-benzimidazolylimino]-3a-(2-chlolophenyl)-6a-phenyl-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole, 5-(2-bezimidazolylimino)-2-[(5-methyl-2-benzimidazolyl)imino]-6a-phenyl-3a-(2-pyridyl)-1,2,3,3a,4,5,6,6a-octahydroimidazo[4,5-d]imidazole bis(trifluoroacetate) salt, 2,5-bis[(1-methyl-2-benzimidazolyl)imino]-3a,6a-diphenyl-1,2,3,3a,4,5,6,6a-octahydromidazo[4,5-d]imidazole, 2,5-bis[(1-methyl-2-benzimidazolyl)imino]-3a,6a-bis(4-methylphenyl)-1,2,3,3a,4,5,6,6a-octahydromidazo[4,5-d]imidazole, and 2,5-bis[(1-methyl-2-benzimidazolyl)imino]-3a,6a-bis(4-bromophenyl)-1,2,3,3a,4,5,6,6a-octahydromidazo[4,5-d]imidazole.

14. A compound of Formula (I) for use as an active therapeutic substance.

15. Use of a compound of Formula (I) in the manufacture of a medicament for use in therapy.

16. A process for preparing a pharmaceutical composition containing a pharmaceutically acceptable carrier or diluent and an effective amount of a compound of the Formula (I) as described in claim 1 and phamaceutically acceptable salts, hydrates, solvates and esters thereof which process comprises bringing the compound of the Formula (I) into association with the pharmaceutically acceptable carrier or diluent.

17. A method of activating the G-CSF receptor which comprises administering non-peptide bifunctional ligand.